Method and system for developing probes for dye normalization of microarray signal-intensity data

Abstract

A method and system for determining a set of dye-normalization microarray probes that consistently hybridize to approximately the same number of target molecules in a wide range of sample solutions. The method of one embodiment of the method of the present invention generates a set of candidate probe molecules. The set of candidate probe molecules are arrayed on one or more replicate microarrays. Sample solutions are made from one or more tissues of one or more species. Microarray-base hybridization assays are conducted by using the replicate microarrays and different sample solutions. A subset of the candidate probe molecules that are functional for the microarray-base hybridization assays are determined.

1 . A method for determining a set of dye-normalization probes, the method comprising: 
 generating a set of candidate probe molecules;    arraying the set of candidate probe molecules on one or more replicate microarrays;    making sample solutions from one or more tissues of one or more species;    conducting microarray-base hybridization assays using the one or more replicate microarrays for each sample solution; and    determining a subset of the candidate probe molecules that are functional for the microarray-based hybridization assays.    
   
   
       2 . The method of  claim 1  wherein generating the set of candidate probe molecules further includes: 
 synthesizing candidate probe molecules having low-complexity, low-entropy nucleotide sequences;    employing candidate probe molecules having low-complexity, low-entropy nucleotide sequences that are complementary to sequenced target molecule; and    employing candidate probe molecules having lengths ranging from about 25 to about 60 or more nucleotides.    
   
   
       3 . The method of  claim 1  wherein the replicate microarrays further includes microarrays having similar feature arrangements.  
   
   
       4 . The method of  claim 3  wherein the replicate microarrays further include one or more features having similar bound candidate probe molecules.  
   
   
       5 . The method of  claim 1  wherein making the sample solutions from one or more tissues of one or more species further includes: 
 isolating a set of target molecules for each tissue;    labeling members of each set of target molecules with identical signal emitting labels; and    labeling each set of target molecules with different signal emitting labels.    
   
   
       6 . The method of  claim 5  wherein isolating the set of target molecules for each tissue further includes amplifying expressed messenger RNA molecules for each tissue.  
   
   
       7 . The method of  claim 5  wherein isolating the set of target molecules for each tissue further includes amplifying complementary DNA molecules for each tissue.  
   
   
       8 . The method of  claim 1  wherein conducting the microarray-base hybridization assays further includes varying one or more of: 
 temperature;    acidity;    alkalinity; and    salinity.    
   
   
       9 . The method of  claim 1  wherein determining the subset of candidate probe molecules that are functional a wide range of microarray-based hybridization assays further comprising: 
 selecting candidate probe molecules that hybridize with target-molecule pairs extracted from different species, tissues, and under varying hybridization conditions;    selecting candidate probe molecules having log ratios within a tolerance interval about zero; and    selecting candidate probe molecules having signal intensities that span the entire intensity distribution range of the microarray-based hybridization assays.    
   
   
       10 . The method of  claim 1  wherein determining the set of candidate probe molecules further includes validating the subset of candidate probe molecules using multiple arrays on a common substrate.  
   
   
       11 . The method of  claim 10  wherein validating the subset of candidate probe molecules further includes arraying approximately 300 of the subset of candidate probe molecules, a quantity of randomly-selected, high-quality biological probes, and a quantity of quality control probes on the multiple arrays on a common substrate.  
   
   
       12 . The method of  claim 11  wherein the randomly-selected, high-quality biological probes further includes probes isolated from tissues of about two or more species.  
   
   
       13 . Transferring results produced by a microarray reader or microarray data processing program employing the method of  claim 1  stored in a computer-readable medium to an intercommunicating entity.  
   
   
       14 . Transferring results produced by a microarray reader or microarray data processing program employing the method of  claim 1  to an intercommunicating entity via electronic signals.  
   
   
       15 . A method comprising forwarding data produced by employing the method of  claim 1  to a remote location.  
   
   
       16 . A method comprising receiving data produced by employing the method of  claim 1  from a remote location.  
   
   
       17 . A system for determining a set of microarray probes, the system comprising: 
 a computer processor;    a communications medium by which microarray data are received by the microarray-data processing system;    a program, stored in the one or more memory components and executed by the computer processor that generates nucleotides sequences for a set of candidate probe molecules; arrays the set of candidate probe molecules on one or more replicate microarrays; and determines a subset of the candidate probe molecules that are functional for two or more tissues of two or more species.    
   
   
       18 . The system of  claim 17  wherein determines the subset of candidate probe molecules further includes determines a log ratio of the one or more sample solutions.